Isolation and characterization of a cDNA for spinach cytosolic ascorbate peroxidase.

Reduced oxygen intermediates are produced in plant tissues from mitochondrial and chloroplastic electron transport pathways and from electron-saturated photosynthetic pigments (Bowler et al., 1992). For instance, the Mehler reaction is a primary source of reduced oxygen intermediates in chloroplasts in which dioxygen is reduced by Fd on the reducing side of PSI to produce .O2-. The dismutation of .O2to H,O, occurs spontaneously or is catalyzed by superoxide dismutases (Neubauer and Yamamoto, 1992). Efficient scavenging of H,O, is critical, since it can damage membranes and deactivate enzymes. In addition, H,O, can react with .O,to produce the highly reactive and dangerous hydroxyl radical (.OH). The reduction of H,O, by ascorbate is catalyzed by APX (EC 1.11.1.11). This enzyme is considered to be an indispensable part of the electron-scavenging pathway (Nakano and Asada, 1987). APX is a heme-containing homodimer found in a11 higher plants (Asada, 1992). Plant APXs include soluble cytosolic isoforms (Mittler and Zilinskas, 1991) and chloroplast-localized isoforms that may be stromal or thylakoid bound (Miyake et al., 1993). We report here the isolation of a full-length cDNA from spinach (Spinaciu oleracea) that encodes a cytosolic APX subunit (Table I). The cDNA is 1102 bp in length with a 750-bp open reading frame, along with 46 bp of 5' and 306 bp of 3' untranslated sequences. The nucleotide sequence of the spinach cytosolic APX open reading frame shows 74% identity with the open reading frame of a cytosolic APX cDNA from pea and 72% identity with one from Arabidopsis. The derived amino acid sequence identity among spinach, pea, and Arabidopsis is 83%.